The rate of musculoskeletal injury differs substantially between men and women, with the incidence of anterior cruciate ligament (ACL) injuries up to 9 times higher in female athletes. These injuries cause short- and long- term health problems, including time lost from school or work, psychological distress, and post-injury joint osteoarthritis. Sex hormones may make a significant contribution to this injury disparity: injury rate differences are not evident until puberty, and injuries occur disproportionately in the first half of the menstrual cycle. While most research has focused on passive components of the musculoskeletal system, arguing that female fluctuating hormone concentrations may change mechanical properties of the ligament itself, a direct link between ligament laxity and ACL injury has not been proven. The goal of this proposal is to test our central hypothesis that changes in sex hormone concentration result in changes to the basic elements of motor control ? at multiple levels, from the musculotendinous unit to motor control circuitry. In our preliminary data, at the muscle level, we have shown that the muscle reflex response is lowest when estradiol concentrations are highest, with no change in anterior knee laxity. At the spinal circuitry level, we have shown that while day-to-day changes in hormone concentration do not modulate spinal motor neuronal excitability, a lower overall level of excitability exists in women who take oral contraceptives. Finally, at the level of the brain, we have shown the excitability of intra-cortical interneuronal circuits is increased when estradiol concentrations are highest. In Aims 1 & 2 of our proposal, daily testing will allow, as never-before, a detailed assessment of the time-specific relationship between hormone concentration and changes at the muscle and spinal level during both relaxed and active states. In Aim 3, daily hormone concentrations will specifically determine each subject's testing day, guiding targeted assessments into the role of fluctuating hormone concentrations on cortical excitability. In Aim 1 we will determine the influence of sex hormone fluctuations on the muscle stretch reflex during active and passive states, and the time-lag between hormone concentration changes and the reflex response. We will use a technically simple assessment that could be implemented in the field. In Aim 2 we will determine the influence of sex hormone fluctuations on spinal motor neuron excitability using H-reflex as a probe and the simultaneous change in the muscle mechanics using muscle twitch response. Aims 1 & 2 will include a focus on the differential role of oral contraceptives. In Aim 3 we will use paired-pulse transcranial magnetic stimulation during active contraction to determine the influence of sex hormone fluctuations on cortical excitability in naturally cycling women. Our findings have the potential to guide future research in injury prevention strategies, including targeted neuromuscular training. This knowledge will enable clinicians to educate female patients on neuromuscular effects of sex hormones during times of high fluctuations, including puberty, pregnancy, and menopause.